Early mixed chimerism-based preemptive immunotherapy in children undergoing allogeneic hematopoietic stem cell transplantation for acute leukemia.

This retrospective analysis comprises 10-year experience with early posttransplant mixed chimerism-based preemptive intervention. Out of 104 patients, 51 received preemptive immunotherapy. Their outcomes were similar to patients achieving full donor chimerism spontaneously. Among patients receiving intervention, 5-year event-free survival was identical in patients with and without pretransplant residual disease, respectively (68% [95% confidence interval (CI) 38-98%] vs. 69% [95% CI 54-85%] log-rank = 0.4). In patients who received preemptive immunotherapy, chimerism status and residual disease prior to transplant were no longer predictors of poor outcome; however, 41% of the patients with residual disease prior to transplant relapsed early and did not benefit from this strategy.

Application of a bromodeoxyuridine-Hoechst/ethidium bromide technique for the analysis of radiation-induced cell cycle delays in asynchronous cell populations.

A flow cytometric technique utilizing the continuous incorporation of bromodeoxyuridine (BrdU) into asynchronous cells to measure radiation-induced cell cycle delay is described. Following the incorporation of the BrdU label the cells are stained with ethidium bromide and the bis-benzimidazole Hoechst 33258. These fluorochromes have differential staining patterns. Hoechst 33258 fluoresces blue and is quenched by BrdU incorporated into cellular DNA during S phase. Ethidium bromide fluoresces red and is not quenched by BrdU. Therefore in cells that are cycling and synthesizing DNA new G1 and G2 compartments are created and this can be used to measure cell cycle delays following ionizing radiation to asynchronous cells. We have used this technique to evaluate two cell lines: a normal diploid human embryo fibroblast cell line MRC 5, which has inducible p53 and shows delays at both G1 and G2 checkpoints, and the human cervix carcinoma cell line HX 156. This cell line has been infected with human papilloma virus (HPV) 16, and therefore has inactivated p53 function and is blocked only at the G2 checkpoint. Using this method, cell cycle-dependent effects relating to the G2 block can be observed. The radiation-induced G2 block differs from that induced by drugs or heating in that cells are blocked in G2 irrespective of the phase of the cell cycle they are treated in. This method allows these different types of G2 block to be quantified.

Initial radiation-induced DNA damage in human tumour cell lines: a correlation with intrinsic cellular radiosensitivity.

The role of the initial DNA double-strand breaks (dsb) as a determinant of cellular radiosensitivity was studied in human breast and bladder cancer cell lines. Cell survival was measured by monolayer colony-forming assay as appropriate and differences in radiosensitivity were seen (alpha-values ranged from 0.12 to 0.54). After pulsed-field gel electrophoresis (PFGE) the initial slopes of dose-response curves were biphasic with a flattening of the curves above 30 Gy. When the frequency of DNA dsb induction was assessed using a mathematical model based on the DNA fragment size distribution into the gel lane, we found a statistically significant relationship between the number of DNA dsb induced and the corresponding alpha-values and fraction surviving after 2Gy (P = 0.0049 and P = 0.0031 respectively). These results support the view that initial damage is a major determinant of cell radiosensitivity.

The usefulness of serum thyroglobulin in the follow-up of differentiated thyroid carcinoma in children.

Serum thyroglobulin has been measured serially in ten children aged 5-17 years presenting with differentiated thyroid carcinoma. At presentation 4 had intra-thyroidal disease, 3 had lymph node metastases, and 3 had lung metastases. During follow-up for a median of 37.0 months (range 21-108) 3 patients have been disease-free, 4 have had a local relapse, and 3 have had persistent disease. Seventy-seven separate serum thyroglobulin measurements have been performed; 36 on and 41 off thyroid replacement therapy. A level of thyroglobulin of less than 5 ng/ml was taken as indicative of absence of disease, and compared against combined clinical examination and 131I scanning. Overall sensitivity of thyroglobulin measurement was 36/37 (97%), and although specificity was 30/40 (75%), this rose to 30/32 (94%) if raised thyroglobulin levels noted within 3 months of 131I therapy in otherwise asymptomatic patients (n = 4) or in subjects with intact thyroid tissue (n = 4) were excluded. Concordance with clinical status was 30/31 (97%) in measurements taken on, and 31/32 (94%) in those taken off, thyroid replacement. These data indicate that thyroglobulin measurement is a sensitive and specific means of detecting residual, recurrent, and metastatic thyroid carcinoma in children.